Revisitation of the phenylisothiocyanate‐derivatives procedure for amino acid determination by HPLC‐UV

Abstract

A revisitation of the well known chromatographic procedure for the determination of amino acids as phenylthiocarbamyl derivatives (PTC) is performed. The method was developed for a microbore column that it is more appropriate to our later aims, the characterization of proteinaceous binders present in microsamples coming from the Cultural Heritage field. Several variables relating to chromatographic aspects were studied such as the pH and temperature of the mobile phase, buffer and modifier (triethylamine) concentrations in mobile phase and the stability of PTC-derivatives in solution. The calibration function was studied in depth. To prevent the heteroscedastic behaviour that it is observed, we used the weighed least squares fitting as the best strategy among other normalizing transformations, such as square root and logarithmic functions. Finally, the proposed method showed results similar to the traditional method in terms of efficiency, runtime, LODs and other characteristics, but with two additional advantages: a lower mobile phase consumption and the possibility of working with a lower sample volume. The usefulness of proposed method is checked against easel painting samples of Pictorial Heritage.

Color and flavor of flaxseed protein hydrolysates Maillard reaction products: effect of cysteine, initial pH, and thermal treatment

“…[10] The quantification was performed by C-18 Inertsil ODS-SP columns and the Waters e2695 Alliance HPLC system (Waters, Milford, MA, USA). [15] Statistical analysis…”

Synthesis of glycylglycine-imprinted silica microspheres through different water-in-oil emulsion techniques

“…The procedure of derivatization was adopted from the work of Calull et al [16] and Checa-Moreno et al [17]. The reaction of derivatization was performed precolumn by adding 50 l of sample to 450 L of a solution of 1% (v/v) PITC in acetonitrile.…”

A validated ultra-performance liquid chromatography with diode array detection coupled to electrospray ionization and triple quadrupole mass spectrometry method to simultaneously quantify taurine, homotaurine, hypotaurine and amino acids in macro- and microalgae

“…Anyway, many derivatizing agents suffer from some limitation; OPA cannot react with secondary amino acids such as proline, FMOC-Cl is fluorescent by itself, and may give rise to disubstituted derivatization products with Tyrosine and Histidine, and Dansyl and Dabsyl chloride reactions proceed very slowly especially with Proline. On the contrary, formation of PITC derivatives is rapid and complete, with both primary and secondary amino acids [31].…”